DNase I sensitive domain of the gene coding for the glycolytic enzyme glyceraldehyde 3-phosphate dehydrogenase

Abstract

A 36-kilobase segment of chicken DNA was cloned, containing the gene coding for glyceraldehyde-3-phosphate dehydrogenase [GAPDH (EC 1.2.1.12)], a glycolytic enzyme which is expressed constitutively in all cell types. Using defined segments of this cloned DNA as probes, the DNase I sensitive domain of the GAPDH natural gene was determined in the hen oviduct. When nuclei isolated from hen oviduct were treated with DNase I under conditions known to preferentially degrade actively transcribed genes (i.e., 15-20% of the DNA rendered perchloric acid soluble), a region of .apprx. 12 kilobase(s) (kb) containing the GAPDH coding sequences and flanking DNA was found to be highly susceptible to digestion by DNase I. Approximately 4 kb downstream from the end of the coding sequences, there was an abrupt transition from the DNase I sensitive or open configuration to the resistance or closed configuration. The chromatin then remained in a closed conformation for at least 10 kb further downstream. On the 5'' side of the gene, the transition from a sensitive to a resistant configuration was located about 4 kb upstream from the gene. Two repeated sequences were localized in the area of DNA that was cloned. One of these is of the CR1 family of middle repetitive elements. It is located about 18 kb 3'' to the gene and as such lies well outside of the DNase I sensitive region which encompasses GAPDH. The other repetitive element is of an uncharacterized family. It is located upstream from the gene and appears to be within a region of transition from the DNase I sensitive to resistant states.